THE UNIVERSITY OF DODOMA

COLLEGE OF HEALTH AND ALLIED SCIENCES

DEPARTMENT OF MEDICAL PARASITOLOGY AND ENTOMOLOGY
TYPE OF WORK: INDIVIDUAL ASSIGNMENT
COURSE NAME: MEDICAL PARASITOLOGY AND ENTOMOLOGY
COURSE CODE: MD 222
COURSE INSTRUCTOR: DR. DAVID MUNIS
DATE OF SUBMISSION: 28 June, 2023
PROGRAMME: DOCTOR OF MEDICINE

NAME REGISTRATION NUMBER

ANGEL LUCAS PINDA T21-03-12664

QUESTION: COMPARE AND CONTRAST HUMAN AFRICAN TRYPANOSOMIASIS AND

SOUTH AMERICAN TRYPANOSOMIASIS.
INTRODUCTION

Trypanosomiasis is the disease of vertebrates caused by different species of protozoa Trypanosoma a

hemoflagellate. The protozoa also cause disease in other animals apart from human beings. The disease in
human beings caused by the protozoa are mainly Human African Trypanosomiasis and South American
Trypanosomiasis. Human African trypanosomiasis is further divided into west and east African
trypanosomiasis.

PROTOZOA MORPHOLOGY

African trypanosomiasis

T. brucei is an elongated spindle-shaped organism. The nucleus is large and centrally located. Kinetoplast
is small and is situated at the posterior. The flagellum starts from the posterior end and runs beyond the
anterior end as free flagellum. The undulating membrane is thrown into 3 to 4 folds. Trypanosoma Brucei
occurs in the vertebrate host as Trypomastigote form. Trypomastigotes show polymorphism. Individual
parasites vary in size and shape in the different phases of their existence.

American trypanosomiasis

T. cruzi occur in two main morphological forms in the human host: trypomastigote and amastigote form.
Trypomastigote measures 20 µ long by 6µ broad; the nucleus is centrally located and the kinetoplast is
large, oval and located posteriorly. The amastigote forms are round and oval bodies and are 2-4 µ in
diameter. Multi-plication of the parasites occurs at this stage only. Trypomastigote forms change to
amastigote forms and the change is reversible.

LIFE CYCLE

All members of the genus Trypanosoma exist at some time in their life cycle, as trypomastigote stage with
an elongated spindle shaped body, central nucleus, a posterior kinetoplast, and long undulating membrane.
Volutin granules are found in cytoplasm. Some trypanosomes such as T. cruzi assume amastigote forms in
vertebrate hosts. In addition to the typical forms, cells with atypical features are frequently found, a
condition known as polymorphism. Trypanosoma pass their life cycle in 2 hosts— vertebrate hosts
(definitive hosts) and insect vectors (intermediate hosts). The vector becomes infective to the vertebrate
host only after an extrinsic incubation period, during which the parasite undergoes development and
multiplication. They then develop differently in their respective vectors.
Human African Trypanosomiasis

Human African Trypanosomiasis is caused by Trypanosoma brucei species (T. brucei rhodiense and T.
brucei gambiense). In the vector, the T. brucei migrate to mouth parts of the vectors, so that infection is
transmitted by vector bite (inoculative transmission).

American trypanosomiasis

The trypomastigotes are transformed into epimastigotes in the midgut, from where they migrate to the
hindgut and multiply. These, in turn, develop into non-dividing metacyclic trypomastigotes (infective
form), which are excreted in faeces (stercorarian transmission) (Martín-Escolano et al., 2022).

TRANSMISION

Human African Trypanosomiasis

For its cyclical transmission, T. brucei depends on tsetse flies. Both sexes are hematophagous and can
transmit trypanosomes. Tsetse are viviparous, and the female deposits a fully developed larva that burrows
into the soil, pupates, and emerges as an adult fly a month later. According to morphological differences
and habitat preference, the thirty one tsetse species and subspecies are classified as forest, riverine or
savannah type (Philippe et al., 2017).

G. fuscipes and G. palpalis (from the palpalis riverine group) are the main vectors of gambiense. For
rhodesiense HAT, the main vectors are G. fuscipes (in Uganda) as well as savannah group species such as
G. morsitans and G. pallidipes (Philippe et al., 2017).

American Trypanosomiasis

In Latin America, T. cruzi parasites are mainly transmitted by the infected faces of blood-sucking triatomine
bugs. These bugs typically live in the cracks of poorly-constructed homes in rural or suburban areas.
Normally they hide during the day and become active at night when they feed on human blood. They usually
bite an exposed area of skin such as the face, and the bug defecates close to the bite. The parasites enter the
body when the person instinctively smears the bug faces into the bite, the eyes, the mouth, or into any skin
break.

T. cruzi can also be transmitted by: food contaminated with T. cruzi through for example the contact with
triatomine bug faces; blood transfusions using blood from infected donors; passage from an infected mother
to her newborn during pregnancy or childbirth; and organ transplants using organs from infected donor’s
laboratory accidents. T. cruzi can also be transmitted to humans via non-vector mechanisms including blood
transfusion and con- genital transmission, which are the main causes of infection in urban areas and non-
endemic countries (Liu & Zhou, 2015).
PATHOLOGY

In African trypanosomiasis, a dramatic increase in IgM is a main feature .In T. b. brucei–infected mice, B
lymphocytes display an aberrant activation phenotype .Antibodies specific to trypanosomes are induced by
several parasite antigens, including variant and invariant glycoprotein epitopes, as well as membrane,
cytoplasmic, and nuclear antigens .

Antibodies directed against trypanosome VSG components appear in sera and their binding to the surface
coat of the trypanosomes induce a decrease in parasitemia, both in the blood and extravascular spaces,
specifically by immune lysis of parasites and their destruction by the Kupffer cells in the liver. Only
heterologous antigenic variants (<0.1%) remain to repopulate the blood and tissues. Parasites are eliminated
due to VSG-specific IgM (appearing at high levels, 3–4 days after infection). During infection, B cell
nonspecific stimulation is enhanced as T-independent B cell responses to the VSG successive parasitemia’s.
In contrast, specific trypanosome B cell response, depending on T cell regulation, is depressed.

Several factors may contribute to this immunosuppression. Macrophages may become unable to present
antigens to T cells (by defects in antigen processing and association of epitopes with MHC Class II) and
produce immunosuppressive factors as nitric oxide (NO), prostaglandins (PG), and cytokines. An increase
in immunosuppressive cytokines, such as IFN-γ and transforming growth factor (TGF)-β, is observed
during infection. However, TGF-β is known to inhibit the production of IL-4, IL-5, and IL-6, the major
cytokines implied in B cell proliferation and differentiation. Autoantibodies directed against components
of CNS myelin have also been reported. They are specific for the major glycosphingolipids of myelin,
the galactocerebrosides, and are detected in the sera of experimentally infected animals and patients from
the Ivory Coast .

Other autoantibodies directed against not yet characterized proteins have been described in HAT patients
as well as antibodies directed at myelin basic protein in experimentally infected animals .Other antibodies
were raised against an epitope containing l-tryptophan, a precursor to the neurotransmitter serotonin, or
recognized some neuronal components of the cytoskeleton, neurofilament proteins. In some cases, these
autoantibodies (antigalactocerebrosides and antineurofilaments) are associated with the neurological stage
of the disease and their detection in sera and CSF could contribute toward defining the neurological
involvement of HAT . In vivo demyelination has been produced by purified antibodies to
galactocerebroside.(Nok, 2009)(Goodwin, 1970)(Kristensson & Bentivoglio, 1999)

In Chagas disease, there are two main hypotheses seeking to explain the mechanisms of pathogenesis in
human Chagas disease. The first of these defends the pivotal role of parasite’s persistence in the host as a
major cause of pathology, while the other postulates that an immune response against self-antigens is
responsible for the tissue damage observed in affected organs of chagasic individuals.

The parasite persistence hypothesis is supported by evidence showing that T. cruzi is present close to, or
within, damaged areas. Although immunohistochemical/histological techniques often fail to reveal
parasites at lesion sites, studies using sensitive techniques such as polymerase chain reaction (PCR) and in
situ hybridization have shown T. cruzi persistence in affected organs. Detected T. cruzi DNA in
endomyocardial biopsies after heart transplantation using PCR. In situ hybridization using human cardiac
tissue provided little evidence for the presence of intact T. cruzi at sites of marked inflammation.
Nevertheless, remnants of both T. cruzi kinetoplast and nuclear DNA were detected.

Furthermore, the transmission of T. cruzi via blood transfusion from chronic chagasic donors and the
observation that there is reactivation of parasitemia in immuno-suppressed patients both support the
assertion that parasites persist in many hosts. Consistent with the parasite persistence hypothesis is the
observation that patients treated with drugs that decrease parasite load display concomitant decrease in
disease severity.

Patients presenting with chronic disease and no heart failure treated with benznidazole exhibited reduced
progression of Chagas disease and increased negative seroconversion. Reduction of electrocardiogram
abnormalities were observed in chagasic patients treated with itraconazole or allopurinol. Thus, it seems
beyond doubt that parasite presence is strongly tied to pathology, since T. cruzi infection is the initial event
responsible for triggering Chagas disease, persists in the host, and because an anti-parasite response is
observed in chronic patients.

On the other hand, the contrast between the severity of the lesions observed during the chronic phase of
Chagas disease and the low parasite load in the blood and tissues of chagasic patients suggests that the
response to the parasite alone is insufficient to account for the observed pathology and that autoreactivity
may contribute to disease aggravation. This idea led to the hypothesis that autoimmune responses take place
during the development of pathology. Favoring this hypothesis is the fact that epitopes of parasite antigens
elicit antibodies that cross react with epitopes of host tissues. Furthermore, it was demonstrated that anti-
parasite host-derived antibodies can mediate cellular reactivity. In addition to the presence of auto reactive
antibodies, many studies demonstrated the existence of auto reactive T-cells in Chagas patients. To this
end, molecular mimicry has been suggested to exist between components of the host and T. cruzi and, thus,
strongly supports the participation of autoimmune reactivity in the pathogenesis.(Dutra et al., 2009)

CLINICAL PRESENTATION

Human African Trypanosomiasis

The clinical manifestations of HAT depend on the parasite subspecies, host response and disease stage.
Variations of virulence and pathogenicity have been attributed to different parasite strains. Both forms of
the disease generally lead to death if untreated, although healthy carriers and self-cure have been described
for gambiense HAT. Rhodesiense HAT is typically acute, progressing to second stage within a few weeks
and to death within 6 months. Gambiense HAT follows a chronic progressive course, with a mean duration
estimated at 3 years, albeit with high interpersonal variability. The disease goes through two stages, a first,
hemolymphatic stage followed by a second, meningoencephalitis stage when trypanosomes cross the blood-
brain barrier and invade the central nervous system (CNS).

First stage gambiense HAT presents predominantly with long-lasting intermittent fever (1 day to 1 week,
separated by intervals of days or months), headache, pruritus, and lymphadenopathy (mainly posterior
cervical, but also possible in the axillar, inguinal and epitrochlear regions). Less frequent features are
hepatosplenomegaly, oedema and endocrine dysfunction (amenorrhea, infertility, miscarriage in women;
reduced libido, impotence in men).

In the second stage, neuropsychiatric disorders add to the first-stage features, while fever becomes less
frequent. The characteristic sleep disorder, which elicited the name sleeping sickness, consists of daytime
somnolence plus sudden overwhelming sleep urges, and nocturnal insomnia. Polysomnographic records
show a disruption of the sleep-wake cycle with frequent, short, sleep-onset REM episodes that occur equally
during day and night. Other neurological signs comprise hyper- or hypo-tonicity, tremor of hands and
fingers and choreiform, athetoid, or oscillatory movements of limbs or trunk, fasciculation, motor
weakness, ataxia, akinesia, and speech disorders. Perioral and cheirooral reflexes are frequently seen.
Mental changes are common, including emotional lability, attention deficit, indifference, apathy, aggressive
behaviour, stereotypic behaviour, dissociative fugue, manic episodes, melancholia, confusion, and
dementia. Neuropsychiatric disorders increase with disease progression 50. Infiltration of endocrine organs
(mainly thyroid and adrenals) and the hypothalamic-hypophysial axis lead to disruption of circadian
rhythms of hormonal secretion including prolactin, renin, growth hormone, and cortisol 47, but generally
do not require specific treatment(Philippe et al., 2017).

American Trypanosomiasis

After exposure to the parasite, a minority of patients will develop an acute syndrome of 4–8 weeks’
duration. Roughly 30–40% of infected patients will subsequently develop the cardiac and/or digestive forms
of chronic CD, usually 10–30 years after the initial exposure.

Chagas heart disease (CHD) is the most common aetiology of cardiomyopathy in Latin America, and a
major cause of cardiovascular death among middle-aged individuals in endemic areas. It manifests as three
major syndromes that often co-exist in the same patient: arrhythmia, heart failure and thromboembolism.
Chronic CD in the gastrointestinal tract leads mostly to megaoesophagus or megacolon. Reactivation
disease can occur in immunocompromised patients producing dermatological manifestations, myo- carditis
and/or meningoencephalitis (Franco-paredes, 1909).

In CHD, arrhythmias are very common and of different types, frequently associated with, and may cause,
palpitations, dizziness, syncope and sudden cardiac death (SCD). Frequent, complex PVCs, including
couplets and runs of NSVT, are a common finding. 24-hour heart failure is often a late manifestation of
CHD. It is usually biventricular with a predominance of left-sided failure at initial stages and of right-sided
failure in more advanced disease. Systemic and pulmonary embolisms arising from mural thrombi in the
cardiac chambers are quite frequent. Sudden cardiac death is the main cause of death in patients with CHD,
accounting for nearly two-thirds of all deaths, followed by refractory heart failure and thromboembolism
(Franco-paredes, 1909) (Chagas Disease (American Trypanosomiasis) Key Facts, 2010).

DIAGNOSIS

Human African Trypanosomiasis

Reliable serodiagnostic tests exist only for gambiense HAT and are based on detection of specific
antibodies. The Card Agglutination Test for Trypanosomiasis (CATT, 58), developed almost 40 years ago,
has been pivotal in the control of gambiense HAT. CATT can be performed with finger prick blood, plasma
or serum and the agglutination reaction is scored visually after 5 minutes. Recently, rapid diagnostic tests
(RDT) for gambiense HAT were developed and introduced in the field: Immune trypanolysis (TL) and
enzyme linked immunosorbent assay (ELISA) are applicable in laboratory conditions on serum, plasma
and dried blood spots (DBS). Their high specificity and sensitivity, their applicability on DBS and their
adaptability to animal specimens make them excellent tools for large scale surveys, post-elimination
monitoring and animal reservoir studies. For T. b. rhodesiense, no field-applicable serodiagnostic test
exists(Philippe et al., 2017)(Mclatchie et al., 2013)

American Trypanosomiasis.

The diagnosis of Chagas disease can be made by observation of the parasite in a blood smear by microscopic
examination. However, a blood smear works well only in the acute phase of infection when parasites are
seen circulating in blood. Diagnosis of chronic Chagas disease is generally based on detection of anti-T.
cruzi IgG antibodies testing with at least two different serologic tests. Determination of CNS involvement
is important to tailor therapy. Evidence of this is defined by either trypanosome seen in CSF or increased
CNS lymphocyte counts (>5 cells/µl). In Rhodesiense HAT, suramin therapy should be commenced
immediately and lumbar puncture should be deferred until blood peristomia has declined (Sudarshi &
Brown, 2015)(Franco-paredes, 1909).
TREATMENT

Human African Trypanosomiasis

Five drugs are used in HAT therapy: pentamidine and suramin to treat first-stage, and melarsoprol,
eflornithine and nifurtimox for second stage disease.

Pentamidine: Pentamidine isethionate is the first-line treatment for first-stage gambiense HAT, and is also
an alternative for rhodesiense HAT, although data on its efficacy against rhodesiense HAT are still limited
55,96 Pentamidine efficacy against ambience HAT (95- 98%) has been stable for decades. It is given
intramuscularly once a day for 7 days, but can also be given in intravenous infusion in saline over 2 h. The
intramuscular injection causes pain and transient swelling. Other adverse events include hypoglycaemia
(5–40%), hypotension, abdominal pain and gastrointestinal problems. Suramin: suramin is used only in
rhodesiense HAT because of the risk of onchocerciasis coinfection in gambiense HAT endemic areas (i.e.
risk of allergic reactions arising from rapid killing of microfilaria), and because pentamidine administration
is simpler. Suramin is administered slowly intravenously. Adverse effects are frequent but mostly mild and
reversible, including pyrexia, nephrotoxicity, peripheral neuropathy, agranulocytosis and
thrombocytopenia.

NECT has higher cure rates (95-98%), lower fatality rates (<1%), less severe adverse events, simpler
administration, and it is believed to avoid drug resistance of the parasite. nifurtimox can only be used to
treat HAT patients off-label, subject to express authorization and responsibility acceptance by national
authorities. The most common treatment-emergent adverse events are abdominal pain, vomiting, and
headache. Eflornithine monotherapy: Eflornithine (α-difluoromethylornithine or DFMO) is given in
monotherapy for gambiense HAT when nifurtimox is unavailable or contraindicated. It is a cytostatic and
trypanostatic drug. Evidence exists that an active immune system is required to achieve cure. Adverse
events are frequent and similar to other cytostatic (including diarrhoea and neutropenia), but eflornithine is
on the whole safer than melarsoprol, with fatality rates below 2%. The main adverse events are fever,
pruritus, hypertension, nausea, vomiting, diarrhoea, abdominal pain, headaches, myelosuppression
(anaemia, leucopoenia, thrombocytopenia), and, more rarely, seizures that are generally isolated and
respond to treatment. Melarsoprol: melarsoprol is restricted to treatment of second- stage rhodesiense
HAT. Close monitoring of patients may allow detection of early signs such as fever and/or headache and
to stop melarsoprol and institute management with dexamethasone and diazepam. Other frequent adverse
reactions include pyrexia, headache, general malaise, gastrointestinal (nausea, vomiting, diarrhoea) and
skin reactions (pruritus); severe complications like exfoliative dermatitis occur in < 1% of cases. Cardiac
failure is common and may be fatal but it may be attributable also to HAT itself.
Treatment in pregnancy Although poorly studied, field experience has accumulated on the management of
pregnant and lactating patients, pentamidine can be given after the first trimester of pregnancy. Nifurtimox,
eflornithine and melarsoprol, all theoretically contraindicated, in practice are given when the mother is in
advanced second stage, and her condition does not permit waiting. If postponing second-stage treatment
until childbirth is judged possible, a pentamidine full course should be administered, principally to prevent
vertical transmission. The benefits and risks must be clearly explained to the patient and her relatives. In
rhodesiense HAT, the acute clinical evolution usually precludes waiting until delivery, and suramin (also
theoretically contraindicated) or melarsoprol are given. New-borns should be examined clinically and
checked for the presence of trypanosomes in the blood. Breastfeeding should continue during HAT
treatment.(Philippe et al., 2017)(Human African Trypanosomiasis (Sleeping Sickness), n.d.)

American Trypanosomiasis

Antitrypanosomal drug treatment is always recommended for acute, congenital and reactivated T. cruzi
infection (most frequently in patients co-infected with HIV) and for chronic T. cruzi infection in children
up to 18 years old. In adults, treatment is currently recommended for those up to 50 years of age with
serologic evidence of T. cruzi infection, based on recent data suggesting that a course of antitrypanosomal
treatment delays progression of cardio- myopathy and improves overall outcomes. There is no evidence
that antitrypanosomal treatment affects the progression of gastrointestinal CD (Franco-paredes, 1909)

Persistence of parasites and the accompanying chronic inflammation is the basis for the pathology in CHD.
Thus, unless demonstrated otherwise by the ongoing randomized controlled trial (BENEFIT),
benznidazole or nifurtimox should be offered to patients <50 years old with presumably long-standing
indeterminate T. cruzi infections or even with mild to moderate disease. Both medicines are fully effective
in curing the disease if given soon after infection at the onset of the acute phase, including the cases of
congenital transmission. The efficacy of both diminishes, however, the longer a person has been infected
and the adverse reactions are more frequent at older age. Benznidazole and nifurtimox should not be taken
by pregnant women or by people with kidney or liver failure. Nifurtimox is also contraindicated for people
with a background of neurological or psychiatric disorders (Franco-paredes, 1909)(Liu & Zhou, 2015).

Medical treatment of CHD is clinically relevant. Amiodarone markedly reduces the frequency and
complexity of ventricular arrhythmias and has been shown to reduce mortality in the only two randomized
trials that included Chagasic patients. Because mild segmental LV wall motion abnormalities are predictors
of deterioration of ventricular function during follow-up, the use of an ACE inhibitor is also highly
recommended. Finally, whether administration of a beta-blocker (to prevent further myocardial damage
and death) and either aspirin or warfarin (to prevent thromboembolism) should be considered, remains an
open and challenging question (Franco-paredes, 1909).
Treatment is also indicated for those in whom infection has been reactivated (for example, due to
immunosuppression), and for patients during the early chronic phase, including girls and women of
childbearing age (before or after pregnancy) to prevent congenital transmission (Liu & Zhou, 2015).

EPIDEMIOLOGY AND DISTRIBUTION

Human African Trypanosomiasis

In the early 20th century, devastating epidemics were probably triggered by the ecological disruptions and
forced population movements brought about by colonialism 10. Since then, the intensity of control efforts
and disease transmission have always been closely linked. In some endemic areas, changes in land use and
climate dramatically reduced tsetse populations and interrupted HAT transmission (Philippe et al., 2017).

American Trypanosomiasis

Chagas disease occurs mainly in Latin America. However, in the past decades it has been increasingly
detected in the United States of America, Canada, many European and some Western Pacific countries.
This is due mainly to population mobility between Latin America and the rest of the world. Less frequently,
it is due to infection through blood transfusion, vertical transmission (from infected mother to child) or
organ donation (Chagas Disease (American Trypanosomiasis) Key Facts, 2010) (Radiological Diagnosis
of Chagas’ Disease (American Trypanosomiasis), 1998)
REFERENCES

Chagas disease (American trypanosomiasis) Key facts. (2010).

http://www.who.int/mediacentre/factsheets/fs340/en/index.html

Dutra, W. O., Menezes, C. A. S., Villani, F. N. A., da Costa, G. C., da Silveira, A. B. M., Reis, D. D. Á.,
& Gollob, K. J. (2009). Cellular and genetic mechanisms involved in the generation of protective
and pathogenic immune responses in human Chagas disease. Memorias Do Instituto Oswaldo Cruz,
104(SUPPL. 1). https://doi.org/10.1590/S0074-02762009000900027

Franco-paredes, C. (1909). American Trypanosomiasis : Chagas Disease. 622.

Goodwin, L. G. (1970). The pathology of African Trypanosomiasis. Transactions of the Royal Society of
Tropical Medicine and Hygiene, 64(6), 797–812. https://doi.org/10.1016/0035-9203(70)90096-9

Human African trypanosomiasis (sleeping sickness). (n.d.). Retrieved June 19, 2023, from
https://www.who.int/data/gho/data/themes/topics/human-african-trypanosomiasis

Kristensson, K., & Bentivoglio, M. (1999). Pathology of African trypanosomiasis. Progress in Human
African Trypanosomiasis, Sleeping Sickness, 157–181. https://doi.org/10.1007/978-2-8178-0857-4_9

Liu, Q., & Zhou, X. (2015). Preventing the transmission of American trypanosomiasis and its spread into
non-endemic countries. Infectious Diseases of Poverty, 1–11. https://doi.org/10.1186/s40249-015-
0092-7

Martín-Escolano, J., Marín, C., Rosales, M. J., Tsaousis, A. D., Medina-Carmona, E., & Martín-Escolano,
R. (2022). An Updated View of the Trypanosoma cruzi Life Cycle: Intervention Points for an
Effective Treatment. ACS Infectious Diseases, 8(6), 1107.
https://doi.org/10.1021/ACSINFECDIS.2C00123

Mclatchie, A. P., Burrell-saward, H., Myburgh, E., Lewis, M. D., Ward, T. H., Mottram, J. C., Croft, S.
L., Kelly, J. M., & Taylor, M. C. (2013). Highly Sensitive In Vivo Imaging of Trypanosoma brucei
Expressing ‘“ Red-Shifted ”’ Luciferase. 7(11), 1–12. https://doi.org/10.1371/journal.pntd.0002571

Nok, A. J. (2009). African Trypanosomiasis. Vaccines for Biodefense and Emerging and Neglected
Diseases, 1255–1273. https://doi.org/10.1016/B978-0-12-369408-9.00062-7

Philippe, B., Giuliano Phd, C., Phd, J. V., & Md, G. (2017). Human African trypanosomiasis.
https://doi.org/10.1016/S0140-6736(17)31510-6

Radiological Diagnosis of Chagas’ Disease (American Trypanosomiasis). (1998). XXXIII(1), 3301.

Sudarshi, D., & Brown, M. (2015). Human African trypanosomiasis in non-endemic countries. Clinical
Medicine, 15(1), 70.